PROJECT SUMMARY Psychomotor activation describes brain commands that guide somatic behaviors. This neurophysical relationship is heavily affected by motivational and emotional states, and is altered in a spectrum of psychiatric illnesses including depression, obsessive-compulsive disorder, autism, ADHD, and Parkinson's disease. Mesolimbic dopamine circuitry, which encompasses dopamine neurons projecting from the ventral tegmental area (VTA) in the midbrain to the nucleus accumbens (NAc) in the ventral striatum, regulates psychomotor behaviors and is broadly implicated in psychiatric disorders. VTA dopamine neurons are heavily constrained by powerful GABAergic inhibition, but how this inhibitory tone influences dopamine-dependent behaviors and psychomotor activity remains unresolved. To define the role of GABA in the modulation of dopamine-dependent behavioral outputs, I am utilizing novel combinatorial genetic and viral strategies to manipulate midbrain GABA and dopamine signaling. I engineered a Cre-dependent, viral-mediated CRISPR/SaCas9 system that allows for cell-type gene knockout. I first validated this technique by targeting Slc6a3, the gene encoding the dopamine transporter (DAT) to produce hyperdopaminergia, and using basal locomotion as a robust and reliable behavioral assay for psychomotor activity. I now propose to use this technique to selectively disinhibit midbrain dopamine or GABA neuron populations by knocking out Gabrg2, the gene encoding the GABAA receptor ?2 subunit (GABAA ?2) in these cell types. The GABAA receptor is the prominent mode of fast, inhibitory neurotransmission within the VTA, and expression of the ?2 subunit is necessary for normal receptor function. Directed elimination of GABAA ?2 will reduce inhibitory tone and therefore allow for characterization of VTA GABA signaling in psychomotor activity. Preliminary experiments suggest disinhibition of VTA dopamine or GABA cells results in psychomotor activation or suppression, respectively. Electrophysiological analysis of intrinsic and extrinsic VTA GABA and dopamine neuron properties will mechanistically verify changes in neurotransmission caused by GABAA ?2 abolition. To further resolve the contribution of GABAergic signaling, I will increase inhibitory tone during conditions of hyperdopaminergic signaling within the VTA through intersectional viral strategies or local drug administration. The results of these experiments will provide insight into how midbrain neural networks organize to generate psychomotor activity, and will help elucidate underlying circuit modifications that result in psychomotor dysfunction in psychiatric illness.